Rhamnolipid-augmented medium enhances growth of <i>Dehalobium chlorocoercia</i> on chloroethenes and polychlorinated biphenyls

ABSTRACT Polychlorinated biphenyls (PCBs) and chlorinated ethenes, such as perchloroethene (PCE), are persistent environmental contaminants that pose significant ecological and health risks. Dehalobium chlorocoercia strain DF-1 can reductively dechlorinate both PCEs and PCBs, offering a sustainable remediation strategy. However, its large-scale cultivation has been limited by dependence on an undefined growth-stimulatory factor produced by Pseudodesulfovibrio sp. (DSV). Filtrates of DSV-autoclaved cultures passed through a 10-kDa cutoff filter enhanced growth-linked dechlorination of DF-1, indicating that the active component is a small-molecular-weight, nonsecretory, thermostable compound. Acid precipitation of the autoclaved DSV extract yielded a glycolipid-enriched fraction that also supported DF-1 growth; in parallel, the model biosurfactant surfactin stimulated DF-1 activity, consistent with a biosurfactant-mediated effect. Liquid chromatography coupled to high-resolution tandem mass spectrometry analysis of the acid-precipitated fraction detected ions and fragmentation patterns consistent with multiple rhamnolipid congeners, including mono- and di-rhamnolipids with C8–C12 acyl chains. Both glycolipid-enriched extract and commercial rhamnolipids substituted for DSV extracts, stimulating DF-1 growth-linked reductive dechlorination of PCE, 2,3,4,5-tetrachlorobiphenyl (PCB 61), and 2,2′,3,3′,4,5,5′,6-Octachlorobiphenyl (PCB 199) in a concentration-dependent manner. Maximal DF-1 growth-linked dechlorination rates were observed at submicellar rhamnolipid concentrations, whereas higher concentrations were inhibitory. Rhamnolipid supplementation reduced growth lag and increased DF-1 biomass and dechlorination rates by solubilizing hydrophobic electron acceptors, thereby increasing their bioavailability in the medium. These findings suggest that rhamnolipid-type biosurfactants in Pseudodesulfovibrio sp. extracts have a role in enhancing the growth of DF-1 and demonstrate that commercial rhamnolipid supplementation can replace undefined coculture extracts, enabling reproducible large-scale cultivation of DF-1 for bioaugmentation of PCB- and PCE-contaminated environments. IMPORTANCE Polychlorinated biphenyls (PCBs) and chlorinated ethenes remain widespread contaminants that are difficult to remove with conventional technologies, creating a need for scalable bioremediation strategies. Dehalobium chlorocoercia DF-1 is one of the few isolates available that can reductively dechlorinate both PCBs and PCE, but its application has been hindered by reliance on undefined extracts from a Pseudodesulfovibrio sp. This study shows that the key stimulatory activity is associated with rhamnolipid-type glycolipid biosurfactants released from Pseudodesulfovibrio sp. cells upon lysis. Glycolipid-enriched extracts and defined commercial rhamnolipids enhance DF-1 growth-linked dechlorination of PCE and PCB congeners in a dose-dependent manner, primarily by increasing the bioavailability of hydrophobic electron acceptors. By replacing complex coculture extracts with a defined biosurfactant supplement, these findings remove a major barrier to reproducible, large-scale cultivation of DF-1 and advance the feasibility of bioaugmentation for PCB- and PCE-contaminated environments.